The invention relates to the design and use of a biological measuring device containing a novel membrane structure.
It is standard practice to treat diabetes mellitus predominantly with insulin injections to compensate for the inability of the pancreas to make insulin to regulate blood glucose levels. The more tightly a person with diabetes is able to regulate his or her blood sugar, the less detrimental the disease is to overall health. The regulation of blood glucose would benefit from a glucose sensing device implanted in the body to monitor blood glucose levels at more frequent intervals than can be done with presently available repeated blood sampling.
A variety of biomedical measuring devices are routinely used by physicians and clinicians to monitor physiological variables such as respiratory rate, blood pressure and temperature. In addition to the repertoire of devices listed above is the enzyme electrode. Enzyme electrodes enable the user to determine the concentration of certain biochemicals rapidly and with considerable accuracy by catalyzing the reaction of a biochemical and a detectable coreactant or producing a product that may be readily sensed by well-known electrodes (e.g. oxygen, H2O2). Currently there are enzyme electrodes that can detect urea, uric acid, glucose, various alcohols, and a number of amino acids when used in certain well-defined situations.
A number of variations of the glucose enzyme electrode have been developed, all based on the same reaction catalyzed by glucose oxidase. 
To accurately measure the amount of glucose present, both oxygen and water must be present in excess. As glucose and oxygen diffuse into an immobilized membrane phase, the glucose reacts with oxygen and water to produce H2O2 (hydrogen peroxide). Glucose is detected electrochemically using the immobilized enzyme glucose oxidase coupled to an oxygen- or hydrogen peroxide-sensitive electrode. The reaction results in a reduction in oxygen and the production of hydrogen peroxide proportional to the concentration of glucose in the sample medium.
The electrode can be polarized cathodically to detect residual oxygen not consumed by the enzymatic process, or polarized anodically to detect the product of the enzyme reaction, hydrogen peroxide. A functional device is composed of at least two detecting electrodes, or at least one detecting electrode and a reference signal source, to sense the concentration of oxygen or hydrogen peroxide in the presence and absence of enzyme reaction. Additionally, the complete device contains an electronic control means for determining the difference in the concentration of the substances of interest. From this difference, the concentration of glucose can be determined.
The enzyme catalase may be included in the oxygen-based system in excess in the immobilized-enzyme phase containing the glucose oxidase to catalyze the following reaction: 
Hence, the overall reaction becomes:
glucose+xc2xdO2xe2x86x92gluconic acid. 
This mixture of immobilized enzymes can be used in the oxygen-based device, but not the peroxide-based device. Catalase prevents the accumulation of hydrogen peroxide which can promote the generation of oxygen free radicals that are detrimental to health.
Glucose measuring devices for testing of glucose levels in vitro based on this reaction have been described previously (e.g. Hicks et al., U.S. Pat. No. 3,542,662) and work satisfactorily as neither oxygen nor water are severely limiting to the reaction when employed in vitro. Additionally, a number of patents have described implantable glucose measuring devices. However, certain such devices for implantation have been limited in their effectiveness due to the relative deficit of oxygen compared to glucose in tissues or the blood stream (1: 50-1000).
Previous devices (e.g. Fisher and Abel) have been designed such that the surface of the device is predominantly permeable to oxygen, but not glucose, and is in contact with the enzyme layer. Glucose reaches the enzyme layer through a minute hole in the oxygen-permeable outer layer that is in alignment with an electrode sensor beneath it. Hydrogen peroxide produced by the enzyme reaction must diffuse directly to the sensing anode or through a porous membrane adjacent to the electrode, but is otherwise substantially confined within the enzyme layer by the oxygen-permeable layer resulting in unavoidable peroxide-mediated enzyme inactivation and reduced sensor lifetime.
The strategy of designing devices with differentially permeable surface areas to limit the amount of glucose entering the device, while maximizing the availability of oxygen to the reaction site, is now common (Gough, U.S. Pat, No. 4,484,987). An example based on device geometry is seen in Gough, U.S. Pat, No. 4,671,288, which describes a cylindrical device permeable to glucose only at the end, and with both the curved surface and end permeable to oxygen. Such a device is placed in an artery or vein to measure blood glucose. In vascular applications, the advantage is direct access to blood glucose, leading to a relatively rapid response. The major disadvantage of vascular implantation is the possibility of eliciting blood clots or vascular wall damage. This device is not ideal for implantation in tissues.
An alternative geometrically restricted device assembly was described in Gough, U.S. Pat, No. 4,650,547. The patent teaches a xe2x80x9cstratifiedxe2x80x9d structure in which the electrode was first overlaid with an enzyme-containing layer, and second with a non-glucose-permeable membrane. The resulting device is permeable to oxygen over the entire surface of the membrane. However, glucose may only reach the enzyme through the xe2x80x9cedgexe2x80x9d of the device in a direction perpendicular to the electrode, thus regulating the ratio of the access of the two reactants to the enzyme.
Devices have been developed for implantation in tissue to overcome potential problems of safely inserting into, and operating sensors within, the circulatory system (e.g. Gough, U.S. Pat. No. 4,671,288); however, their accuracy may be limited by the lower availability of oxygen in tissues. The device membrane is a combination of glucose-permeable area and oxygen-permeable domains. The ratio of the oxygen-permeable areas to the glucose-permeable areas is somewhat limited due to the design.
To avoid geometric restrictions on devices, membranes that are variably permeable to oxygen and glucose have been developed (Allen, U.S. Pat. No. 5,322,063). Membrane compositions are taught in which the relative permeability of oxygen and glucose are manipulated by altering the water content of a polymeric formulation. The disadvantages of such a membrane may include sensitivity of the membrane performance to variables during manufacture and that regions of oxygen permeability may not be focused over electrodes within the device.
An alternative strategy to device construction is to incorporate an enzyme-containing membrane that is hydrophilic and also contains small hydrophobic domains to increase gas solubility, giving rise to differential permeability of the polar and gaseous reactants (e.g. Gough, U.S. Pat. Nos. 4,484,987 and 4,890,620). Such membranes readily allow for the diffusion of small apolar molecules, such as oxygen, while limiting the diffusion of larger polar molecules, such as glucose. The disadvantage is that the amount of hydrophobic polymer phase must be relatively large to allow for adequate oxygen permeability, thereby reducing the hydrophilic volume available for enzyme inclusion sufficient to counter inactivation during long-term operation.
Schulman et al. (U.S. Pat. No. 5,660,163) teach a device with a silicone rubber membrane containing at least one xe2x80x9cpocketxe2x80x9d filled with glucose oxidase in a gelatinous conductive solution located over a first working electrode. In a preferred embodiment, the length of the xe2x80x9cpocketxe2x80x9d is approximately 3 times its thickness to optimize the linearity between current and the glucose concentration measurement. Because the long axis of the xe2x80x9cpocketxe2x80x9d is oriented parallel to the electrode surface, this design may be less amenable to miniaturization for tissue implantation.
The invention is the design and use of a biological measuring device for implantation into an individual or for use in an external environment. The device contains an enzyme electrode to detect the coreactant or product (e.g. oxygen, H2O2, respectively) of an enzymatic reaction catalyzed by an oxidase (e.g. glucose oxidase, lactate oxidase, cholesterol oxidase) of the biological molecule of interest (e.g. glucose, lactate, cholesterol) with a limiting reagent or coreactant (e.g. oxygen). The device contains a differentially permeable membrane that limits the access of the biological molecule of interest, which is present in the device""s environment at a relatively high concentration as compared to the coreactant, to the enzyme. (Expected ratios of biological molecule to coreactant concentrations (e.g. glucose concentration to oxygen concentration) in biological samples or environments may be expected to range up to 10:1 and beyond, expressed in units of mg/dl/mmHg.) Thus, the biological molecule becomes the limiting reagent in a critical zone within the enzyme-containing region of the membrane, allowing for its quantification by assaying the amount of product produced or the amount of unconsumed coreactant by means of an associated sensor or electrode, responsive to the coreactant or product.
The membrane is composed of a continuous or nearly continuous restricted-permeability membrane body, permeable to oxygen and essentially impermeable to larger biological molecules (e.g. glucose, lactate, cholesterol), and discrete hydrophilic regions, permeable to both biological molecules and oxygen (FIG. 1). The reactants diffuse from the environment into the device through a single surface of the device. The size, density, shape, and number of hydrophilic regions may be varied depending upon the bodily fluid, tissue, or environment into which the device is implanted or depending upon the choice of the associated sensor. As opposed to prior membranes which have restricted-permeability and hydrophilic surfaces at restricted locations on the device defined by device shape, or other sensors covered in membranes whose differential oxygen- and biological molecule-permeability is continuous, the location, number, shape, and size of the oxygen- and biological molecule-permeable regions may be modified to optimize the performance of the sensor.
The invention is a biological measuring device containing the composite membrane of the invention. The membrane of the invention can be optimized for detection of a number of biochemicals with a single or a plurality of detecting electrodes. Electrodes may be linked in any of a number of ways well known to those skilled in the art (e.g. Sargent and Gough, 1991, herein incorporated by reference). The size, shape, number, and location of the hydrophilic regions can be varied to deliver the appropriate ranges of the biological molecule and oxygen to the enzyme such that a detectable amount of product or consumed coreactant reaches the associated sensor.
The invention is a method to specify the optimal ratio of restricted-permeability membrane body to hydrophilic regions in the membrane, and to determine the optimal shape and arrangement of the hydrophilic regions in the membrane such that the concentrations of the reactants in the critical zone are limited by diffusion. Using the method of the invention, the sensor can be optimized for different reactions and enzymes for use in different tissues, bodily fluids or in an external sensor.
The invention is the use of the biological measuring device to monitor the level of a biological molecule, either by implantation in an individual or by use of the device in an external environment. In a preferred embodiment, the device is used to monitor glucose levels in an individual with diabetes.